Fetal bovine serum: are cell cultures cruelty free?
By Carlo Jochems, M.Sc. ¨C [email protected]
Cell culture techniques represent a way of performing scientific and medical research without the use of animals who are hampered with. At least that is the view many people hold. However, several steps in the setting up of functional cell cultures may involve the use of animals and hence imply welfare aspects. Consequentially, what is considered to be the major alternative technique in animal experimentation ¨C cell culture ¨C itself implies animal welfare aspects as well. In this paper the focus is on the production method of fetal bovine serum (FBS) and its welfare and ethical implications.
To culture cells, tissues or organs in a plastic flask, one has to arrange proper growing circumstances for these cells. This is done by adding so called culture medium to the cells, usually combined with fetal bovine serum (serum is blood without cells and clotting factors). Fetal bovine serum contains many substances which are needed by cultured cells to grow and live properly. FBS is the serum of first choice for culturing cells, tissues and organs in vitro due to its broad applicability: nearly every cell type thrives with it. It is being used extensively both in research and (the life science-) industry, making fetal bovine serum a commercially interesting product. FBS is the serum of an unborn bovine i.e. a bovine fetus. Let's take a closer look on its whereabouts before it is delivered in sterile plastic bottles into the cell culture laboratory.
Fetal bovine serum is obtained in those areas of the world where bovines are kept in an extensive manner (e.g. continent of America, Australia, Southern Africa, CIS). That is, herds in which cows and bulls roam freely amongst eachother. Bulls are added to herds of cows to help maintaining the herd together, just like dogs keep a flock of sheep together. As a result however, several cows will be pregnant by the time the herd is sent to slaughter (the herd is sent in its entirety to slaughter when the average age of the cows is around two years). This is common practice, whether or not a cow is pregnant is simply not an issue! So, once at the slaughterhouse, all cows are slaughtered, including the pregnant ones. During removal of the internal organs of any cow on the slaughterline, a fetus may show up. Then, if the slaughterhouse has the proper facilities, the entire reproductive tract including the fetus is cut quickly out of the carcass of the mother cow and brought to a more septic area (called: calf processing area) where the following procedure is followed to obtain its blood.
First, the fetus is cut out of the uterus. The umbilical cord is tied off, the fetus is cleaned from amniotic fluid and desinfected. Then, a large diameter needle is inserted through the skin and between the ribs, directly into the beating heart of the unanaesthesized fetus. The blood is commonly extracted under vacuum into a sterile blood collection bag via a tube connecting the two. In the absence of a vacuum pomp, fetal blood may be obtained by means of gravity. Once the blood has been obtained, it is allowed to clot at low temperature, after which process the clotted substance is separated from the serum by refrigerated centrifugation. Afterwards, the ¡®empty¡¯ fetus is destructed to assure it will not be used for human consumption.
The heart of the fetus must function in order to obtain an adequate (read: commercially satisfactorily) amount of fetal blood for FBS production. If the heart functions, the fetus is - by definition - alive. But it is not receiving any form of anesthesia prior to being exposed to a cardiac puncture, which represents a problem because it is a very painful procedure in animals after birth. The last ten to fifteen years more and more scientific data is piling up showing that the fetuses of mammals (in particular those of the species whose newborns are relatively well-developed at birth, like bovines, horses, guinea pigs, sheep, goats, pigs) can experience pain or discomfort well before birth. In a recent guideline on the humane euhanasia of experimental animals, it is said that such animals could experience pain or discomfort as early as 30% gestation time. For a bovine fetus this is as from 3 months as the total gestation period is 9 months. Bovine fetuses used for FBS harvest must at least be 3 months of age (otherwise they are simply too small), but commonly they are of 6 months of age or older. So, all bovine fetuses used for FBS production are capable of sensing pain, yet they are never anaesthesized! What makes it even worse, is the finding that mammal fetuses are not just able to feel pain from a certain timing in their development, they are even more susceptible to pain than adults. After birth, this ¡®over¡¯-susceptibility decreases to adult values as the newborn grows older and ¡®dampening¡¯ systems in the neural pathways become active.
Apart from the difficulties in themselves of assessing possible suffering in a fetus, in this case this is also hampered by the lack of oxygen in the fetus resulting from the few minutes between killing of the mother cow and the performance of a cardiac puncture (lack of oxygen is a common means of causing unconsciousness as part of anesthesia) but exactly ¡®how¡¯ unconscious a bovine fetus is at the moment of puncture has never been scientifically investigated - as far as I know at least. Moreover, again showing the differences in physiology between mature and fetal mammals, mammal fetuses are highly resistant to lack of oxygen as compared to adults of the same species. A fetus survives lack complete lack of oxygen much longer then does its adult counterpart. In normal laboratory settings, this would imply that a scientist wishing to perform a cardiac puncture on a bovine fetus for serum harvest using the procedure described above, should consider anaesthesizing the fetus somehow before the puncture. This a applied ethics, as it is normal that scientists err on the side of caution wherever possible animal suffering may be involved, not on the side of ignorance, disinterest or the-easiest-way.
This represents an ethical problem, which sheds another light on its use in so-called “®alternative methods” replacing “®direct” animal experiments. According to the current animal experimentation legislation of the UK, the procedure described above would be considered an animal experiment (all fetuses are covered by that law as from 50% gestation period). A more open FBS-seller says that bovine fetuses would be or have been bred specifically for the purpose of harvesting their blood in some central European countries. Apparently, there is something as a contradictio in vitro when it comes to the use of FBS in the culturing of cells. Per year around 500,000 liters of FBS are obtained from > 1,000,000 bovine fetuses worldwide.
With the exception of a few, commercial suppliers are almost all very reserved to give information on any aspect of FBS production: now you know why! Finally, most people oppose to animal experiments (or: causing harm to animals) for the purpose of e.g. make-up and toiletry research. Blood sampling is considered a procedure in most Western countries; should this be allowed for indirect make up research if their serum is used for in vitro experiments? Should the unborn bovine fetus suffer for the same purpose? Or, does the fact that it is currently unknown to what extent the bovine fetus suffers, mean the fetus should not be anaesthesized?
Concerns over the periodic fluctuations in cost and availability of fetal bovine serum, its quality (contaminations, support of cell growth, smuggling), imprecise composition (batch-to-batch variation, presence of substances with unknown function) and related ethical issue stimulate the creative quest for fetal bovine serum alternatives. Substitutes include 100% synthetic media (cell type specific = expensive to develop), reduced-serum media (broader applicability then synthetic media), alternative post-natal mammalian serum substitutes like new-born calf, horse, adult bovine and donor bovine serum (more endotoxins and antibodies present, in particular in new-bron calf serum; still animal welfare aspects), fetal horse / pig serum (other composition than FBS, same ethical problem). A colostrum-based alternative has been withdrawn from the market. It seems fair to say that 100% synthetic medium is the best alternative currently at hand. The cell type cultured determines the optimum serum (substitute) and its concentration.
Altogether, what should be kept in mind is (1) fetuses of many mammal species can feel pain already long before the moment of birth, (2) the use of fetal bovine serum represents currently an ethical problem because it is not clear to what extent the unborn cow feels pain from the heart puncture, (3) the use of other animal sera (like donor sera) also implies certain animal welfare aspects becoming related to cell culture work, as does (4) the killing of animals for the purpose of getting cells to be cultured in vitro. The overall conclusion is that cell culture work does not free us from animal welfare considerations as taken for granted by many!
For more information on the issue of fetal bovine serum and its alternatives, you may wish to visit the following site: http://www.nca-nl.org , option <Documents>, which contains the summary of the M.Sc.-thesis report on livestock blood sera production methods and related ethical implications, as well as a downloadable version of the complete report. A scientific article on this issue has been published in the journal Alternatives To Laboratory Animals (30 (2), 2002). Because this is worthwhile in itself in terms of animal welfare but also to inventarise the ethical consequences for cell culture techniques as alternatives to animal experiments.
Carlo Jochems, M.Sc. ¨C [email protected]
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